Storage-driven microbial and quality dynamics of brewer’s spent grain for food valorization

• Storage temperature and duration govern BSG stability and microbial succession. • Pseudomonas dominated at 4°C, while Bacillus spp. dominated at 20°C and 35°C. • High temperature storage accelerated nutrient loss and enhanced bioactive compound. • Microbial shifts explained 50% biochemical and 49.6% volatile changes. • Spoilage at 20–35°C is driven by Bacillus -related genera. Brewer’s spent grain (BSG), a nutrient-rich by-product of brewing, holds significant potential for food applications; however, its high moisture content promotes rapid spoilage, limiting its utilization. The mechanistic links between microbial succession and the dynamics of biochemical and volatile compounds during storage remain poorly understood. This study provides an integrated analysis of microbial succession (16S rRNA and ITS2 sequencing), biochemical transformations, and volatile dynamics of BSG stored at 4°C, 20°C, and 35°C. The fresh BSG microbiota was dominated by Proteobacteria, Firmicutes, Deinococcota, Actinobacteriota, and Ascomycota. Storage temperature, duration, and their interaction explained variation in bacterial composition (R² = 21.9%, 19.9%, and 41.1%, respectively), while fungal communities remained relatively stable. At 4°C, Pseudomonas remained dominant, whereas warmer conditions (20–35°C) favored Bacillus -related genera (75–90% relative abundance). These microbial shifts significantly altered biochemical composition, driving macronutrient degradation and increases in organic acids (3.24-fold), total phenolics (3.2-fold), antioxidant activity (8.7-fold), and an increase in pH (6.5 to 8.8). Bacterial community dynamics explained 50.0% of biochemical variation and 49.6% of volatile profiles, highlighting strong microbiome–metabolite interactions. Refrigeration (4°C) slowed degradation and preserved nutrients; however, late-stage relative enrichment of Pseudomonas indicates that spoilage was delayed rather than prevented. Sensory deterioration, including color change, stickiness, and off-odor development, was pronounced at 20 and 35°C. Overall, temperature-driven microbial succession is the primary driver of BSG quality loss, providing insights to optimize storage, extend shelf life, and ensure safety in food valorisation.